title: Characterization of the glycosylphosphatidylinositol-anchor signal sequence of human Cryptic with a hydrophilic extension.
abstract: Epidermal Growth Factor-Cripto-1/FRL-1/Cryptic (EGF-CFC) proteins, including human Cripto-1 (hCFC2/hCR-1) and human Cryptic (hCFC1), are membrane-associated Nodal co-receptors, which have critical roles in vertebrate development. Most of the EGF-CFC proteins have been experimentally proven or predicted to be glycosylphosphatidylinositol (GPI)-anchored proteins. However, unlike other EGF-CFC proteins, hCFC1 does not exhibit a typical GPI-signal sequence, containing a 32-amino acid hydrophilic extension in its COOH-terminal end. Here we experimentally demonstrate that the COOH-terminal sequence of hCFC1 functions as a GPI-anchoring signal. Moreover, addition of a hydrophilic epitope tag of 55-amino acids (V5-His) after the GPI signal of hCR-1 interfered with generation of a GPI-anchored form of hCR-1. In contrast, addition of the same epitope tag to the end of GPI signal of hCFC1 did not affect the GPI-attachment of hCFC1. The COOH-terminal signal of hCFC1 could produce two different forms of the protein; a GPI-anchored form and an unprocessed form which was more prone to be secreted into the conditioned medium. The hydrophilic extension of hCFC1 negatively regulates the activity of hCFC1 as a Nodal co-receptor. These results demonstrate the presence of endogenous GPI-signal sequence with a hydrophilic extension, which can generate both GPI-anchored and soluble forms of the protein.
authors.affiliation: Tumor Growth Factor Section, Mammary Biology and Tumorigenesis Laboratory, Center for Cancer Research, National Cancer Institute, National Institutes of Health Bethesda, MD 20892-4254, USA.
authors.forename: Kazuhide
authors.initials: K
authors.lastname: Watanabe
authors.forename: Tadahiro
authors.initials: T
authors.lastname: Nagaoka
authors.forename: Luigi
authors.initials: L
authors.lastname: Strizzi
authors.forename: Mario
authors.initials: M
authors.lastname: Mancino
authors.forename: Monica
authors.initials: M
authors.lastname: Gonzales
authors.forename: Caterina
authors.initials: C
authors.lastname: Bianco
authors.forename: David S
authors.initials: DS
authors.lastname: Salomon
mesh_terms: D000595:Amino Acid Sequence; D002460:Cell Line; D004815:Epidermal Growth Factor / Q000235:genetics / Q000378:metabolism*; D058851:GPI-Linked Proteins; D017930:Genes, Reporter; D017261:Glycosylphosphatidylinositols / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D007150:Immunohistochemistry; D036341:Intercellular Signaling Peptides and Proteins / Q000235:genetics / Q000378:metabolism*; D007668:Kidney / Q000166:cytology; D008156:Luciferases / Q000378:metabolism; D008562:Membrane Glycoproteins / Q000235:genetics / Q000378:metabolism*; D008969:Molecular Sequence Data; D009363:Neoplasm Proteins / Q000235:genetics / Q000378:metabolism*; D055457:Nodal Protein / Q000235:genetics / Q000378:metabolism*; D021382:Protein Sorting Signals / Q000235:genetics / Q000502:physiology*; D014162:Transfection
chemical_list: C491268:CFC1 protein, human; D058851:GPI-Linked Proteins; D017261:Glycosylphosphatidylinositols; D036341:Intercellular Signaling Peptides and Proteins; D008562:Membrane Glycoproteins; D009363:Neoplasm Proteins; D055457:Nodal Protein; D021382:Protein Sorting Signals; C070636:TDGF1 protein, human; D004815:Epidermal Growth Factor; D008156:Luciferases